Single-cell dissection of persistent tumor antigens in non-responders reveals opportunities for TAA-targeted vaccination after neoadjuvant therapy in esophageal squamous cell carcinoma

Background Neoadjuvant immune checkpoint blockade (ICB) combined with chemotherapy has demonstrated higher response rates than chemotherapy alone in resectable esophageal squamous cell carcinoma (ESCC). Although tumor microenvironment (TME) features associated with treatment response have been studied in the contexts of both chemotherapy and immunotherapy, direct comparisons of these characteristics and their implications for novel therapeutic development remain largely unexplored. Methods Paired pretreatment and post-treatment tumor samples were prospectively collected from 55 patients with locally advanced ESCC enrolled in a multicenter, phase 3 clinical trial (ChiCTR2000040034). Patients were randomized to receive neoadjuvant chemotherapy (paclitaxel with cisplatin, TP; n=14) or its combination with PD-1 blockade (camrelizumab, albumin-bound paclitaxel and cisplatin/paclitaxel and cisplatin, Cam+nab-TP/TP; n=41), followed by surgical resection. Pathological response was defined based on Mandard’s Tumor Regression Grade scoring system. Changes in immune cell populations and tumor-associated antigens (TAAs) within the TME in relation to response were characterized using single-cell RNA and T cell receptor sequencing. Results Both chemotherapy and chemoimmunotherapy responders exhibited shared temporal dynamics including dendritic cell remodeling, cytotoxic CD8 + T cell contraction, and memory T cell expansion, with chemoimmunotherapy uniquely suppressing the clonal expansion of GZMB + TIGIT + T cells. In contrast, non-responders failed to elicit effective antitumor immunity and displayed persistent expression of targetable TAAs, fully preserved HLA machinery, and clonal expansion of dysfunctional GZMB + TIGIT + T cells. Conclusions Our study identifies key immune contributors correlated with response to neoadjuvant therapies and a panel of TAAs in non-responders. These findings support the development of TAA-targeted therapeutic vaccines and combination strategies incorporating ICB to overcome resistance in non-responders.